Tissue interactions in normal mammary gland are important regulatory mechanisms in homeostatic maintenance of the gland. In the mouse, both normal and preneoplastic mammary epithelia are stroma-dependent, that is, they only grow in mammary fatpads in vivo, but they are inhibited by normal epithelium. Thus, for growth, normal and preneoplastic cells must be injected into mammary glands free of glandular elements. Mammary tumor cells are no longer stroma-dependent but are still stroma- responsive since growth is stimulated by stroma in mammary fatpads versus subcutaneous sites. The interaction with normal mammary epithelium is, on the other hand, the opposite of preneoplastic epithelium since growth of neoplastic cells is stimulated rather than inhibited in the presence of normal mammary epithelium. To define the role of intraepithelial and stromal-epithelial interactions in breast tissue homeostasis and subsequent alterations during progression, we have developed an assay for diffusible growth factors in which both producer and responder cells grow in a 3-dimensional array in collagen gel matrix. The effect of normal mammary cells on neoplastic and preneoplastic cell growth in this assay accurately reflects in vivo events; that is, both normal mammary epithelium and normal mammary stroma stimulate growth of mammary tumor cells but not of normal mammary epithelium or preneoplastic HAN (hyperplastic alveolar nodules) cells. However, in parallel experiments using monolayer cultures, neoplastic cells are inhibited by normal mammary cells in vitro. Thus, matrix, cell shape, and/or multicellular architecture are critical to homeostatic interactions. Results with the collagen gel assay also indicate that mammary tumor cells are stimulated by a mammary stromal fibroblast line but that preneoplastic cells require additional stromal elements for stimulation to occur. We have developed a second generation of the collagen assay which allows detection of contract-dependent interactions as well as those mediated by soluble factors and propose to define stromal elements responsible for stimulating growth of mammary epithelial cells. Furthermore, the mammary stroma in situ is a complex structure of fibroblasts, lipocytes, vasculature, and infiltrating lymphokines in the growth regulation of mammary epithelial cells at different stages of progression will be the focus of this proposal.
